The present invention relates to a biological molecules detection device having increased detection rate, and to a method for quick detection of biological molecules.
Typical procedures for analyzing biological materials, such as nucleic acid, involve a variety of operations starting from raw material. These operations may include various degrees of cell purification, lysis, amplification or purification, and analysis of the resulting amplified or purified product.
As an example, in DNA-based blood tests the samples are often purified by filtration, centrifugation or by electrophoresis so as to eliminate all the non-nucleated cells. Then, the remaining white blood cells are lysed using chemical, thermal or biochemical means in order to liberate the DNA to be analyzed.
Next, the DNA is denatured by thermal, biochemical or chemical processes and amplified by an amplification reaction, such as PCR (polymerase chain reaction), LCR (ligase chain reaction), SDA (strand displacement amplification), TMA (transcription-mediated amplification), RCA (rolling circle amplification), and the like. The amplification step allows the operator to avoid purification of the DNA being studied because the amplified product greatly exceeds the starting DNA in the sample.
The procedures are similar if RNA is to be analyzed, but more emphasis is placed on purification or other means to protect the labile RNA molecule. RNA is usually copied into DNA (cDNA) and then the analysis proceeds as described for DNA.
Finally, the amplification product undergoes some type of analysis, usually based on sequence or size or some combination thereof. In an analysis by hybridization, for example, the amplified DNA is passed over a plurality of detectors made up of individual oligonucleotide probe fragments (probes) that are anchored, for example, on electrodes. If the amplified DNA strands are complementary to the probes, stable bonds will be formed between them and the hybridized probes can be read by observation using a wide variety of means, including optical, electrical, mechanical, magnetic or thermal means.
Other biological molecules are analyzed in a similar way, but typically molecule purification is substituted for amplification and detection methods vary according to the molecule being detected. For example, a common diagnostic involves the detection of a specific protein by binding to its antibody or by a specific enzymatic reaction. Lipids, carbohydrates, drugs and small molecules from biological fluids are processed in similar ways.
In known microfluidic devices, the detectors are generally arranged in an array within a detection chamber. The probes are anchored in predetermined array locations to respective basements, which may be made either of conductive or insulating material. For example, the basements frequently comprise metal electrodes deposited on a bottom wall of the detection chamber. As an alternative, the probes are fixed to doped conductive regions in a semiconductor layer, such as a silicon substrate or an epitaxial layer. Moreover, conducting basements are often coated with a dielectric passivation layer for providing electrical insulation and protection against undesired chemical interactions between the electrodes and the biological sample in the detection chamber. Basements may be regions of a non-conducting material as well, such as silicon dioxide, silicon carbide, silicon nitride, undoped silicon, gels, polymers, and similar materials.
Several known techniques are used for anchoring the probes to the respective basements, such as spotting, chemical grafting, covalent binding, surface adsorption, or electrochemical methods.
The detection chambers presently exploited in microfluidic devices for biological analysis have some limitations. In order for hybridization to occur, for example, target DNA needs to be in the vicinity of its complementary probe. However, target DNA moves according to concentration gradients and random thermal motion. Hence, the DNA moves very slowly. Interactions between a probe and its corresponding target are accordingly quite unlikely and hybridization takes a very long time. Disadvantageously, the step of detection usually takes at least 20-30 minutes and, more frequently, about 1-2 hours or even more.
The aim of the present invention is to provide a detection device and method that are free from the above described drawbacks.